This mentored clinical scientist development award (K08) proposal describes the 5 year training program for Dr. Matthew J. Schuchert. This proposal builds upon the Candidate's strengths and prior research skills and takes advantage of the guidance and resources of the research scientists at the University of Pittsburgh. Through the mentorship of Dr. Angus Thomson, as well as a structured didactic component, the Candidate's ability to perform hypothesis-driven research will be advanced to a fully-independent surgeon scientist. The broad, long term objective of this project is to evaluate the mechanism(s) by which CD8+ marrow- derived subsets (CD8+/TCR-) enhance (facilitate) hematopoietic stem cell engraftment across allogeneic barriers In mice, resulting In the development of multllineage chlmerism and donor-specific transplantation tolerance without GVHD. Recent studies have demonstrated that cells resembling both plasmacytold pDCs (CD8+/CD11c+/B220+/CD11b-) and developing T cells (pTCs: CD8+/CD3+) exist within the FC population, which individually fail to achieve the complete facilitation effect. Therefore, we hypothesize that facilitation of allogeneic hematopoietic stem cell engraftment Is enhanced by the co-adminlstratlon of discrete CD8+ pDC and pTC progenitors working in concert to augment their tolerogenic effect. Specific Aim 1: Evaluate the functional contributions of individual CD8+/TCR- marrow-derived subsets. - Co-administration of pDC and pTC subsets will be assessed in vivo to examine facilitating potential. Specific Aim 2: Assess the cell trafficking and differentiation patterns of pDCs and pTCs. - EGFP and luclferase labelled pDC and pTC will be studied following allogeneic transplantation. Specific Aim 3: Evaluate potential mechanisms of FC function in vitro and in vivo. - Will evaluate potential mechanisms related to co-stimulation, cytokine polarity and Treg induction Specific Aim 4: Identification and characterization of facilitating subsets In human marrow. - Will characterize CD8+/TCR- subsets In human marrow, and assess facilitating potential in NOD-scid mice. Data from these studies will shed important Insights on cell-based therapies for tolerance induction. RELEVANCE (See instructions): A better understanding of pDC and pTC interactions will aid the development of innovative purified cell component therapeutic strategies for the treatment of hematopoietic malignancies, as well as for the support of human stem cells in a wide variety of clinical settings (hemaglobinopathies, immunodeficiencies, tissue regeneration, and ischemic heart failure). Such knowledge would afford new hope in thoracic organ transplant recipients and in patients suffering from a wide range of congenital and acquired hematologic disorders.